Shiga toxin-producing Escherichia coli, including O157:H7, are emerging pathogens of major importance. Bacterial colonization is associated with intestinal disturbances; however, the severe, potentially fatal symptoms of bloody diarrhea (hemorrhagic colitis) and destruction of the kidney leading to hemolytic uremic syndrome (HUS) are due to a bacterial toxin, Shiga toxin. The genes for Shiga toxin are encoded by a temperate bacteriophage in the late gene operon. Shiga toxin is expressed when the phage lytic cycle is induced, along with the genes necessary for production of viral particles and bacterial lysis. Phage-mediated bacterial lysis is necessary for toxin secretion. The viral particles produced during infection with the pathogenic O157:H7 can infect the nonpathogenic intestinal E. coli. When this occurs, the previously harmless E. coli will produce Shiga toxin and amplify the pathogenic process. Our preliminary data suggest that the Shiga toxin production by intestinal flora can be substantial and that severe, possibly life-threatening disease is more likely to occur in individuals whose intestinal flora can be infected by the Shiga toxin phage. In contrast, individuals with intestinal flora that are resistant to the Shiga toxin phage will be protected from severe disease. [unreadable] [unreadable] Currently, it is difficult to prevent disease by E. coli O157:H7 since the infectious dose is very low, and antibiotic treatment, instead of being beneficial, can enhance progression to fatal disease. We propose to develop methods to convert the intestinal flora to phage-resistance as a therapeutic approach to controlling disease caused by Shiga toxin-producing pathogens. This is especially important since there is no treatment, other than supportive care, once disease has developed. The specific aims of this application are to: 1) Characterize Shiga toxin production by clinical isolates of O157:H7 in the presence of susceptible or resistant human intestinal E. coli; and 2) Develop methods to convert intestinal flora to phage resistance using a mouse model of disease. [unreadable] [unreadable]